Propylene is an important basic raw material for synthetic resins such as polypropylene or petrochemical products, and propylene widely is used in bumpers for cars, food containers, films, medical instruments, and the like.
Isopropyl alcohol produced from a plant-derived raw material can be converted to propylene through a dehydration process. Therefore, isopropyl alcohol is expected to be useful as a raw material for carbon-neutral propylene. Considering the current situation in which Kyoto Protocol mandates reduction of total amount of greenhouse gas emitted from developed countries by 5% during the period of 2008 to 2012 as compared to that in 1990, propylene, which is carbon-neutral, is extremely important from the viewpoint of global environment due to its versatility.
Microorganisms that assimilate a plant-derived raw material to produce isopropyl alcohol are already known.
For example, WO 2009/008377 discloses that isopropyl alcohol is produced using an Escherichia coli modified to produce isopropyl alcohol from glucose as a raw material while a semibatch cultivation with sequential addition of a substrate solution is carried out. It is described that this isopropyl alcohol-producing Escherichia coli has excellent properties as a biocatalyst for industrial production because of its high isopropyl alcohol selectivity.
In order to produce isopropyl alcohol at the industrial level using a culture method, effective production of isopropyl alcohol through long-term continuous cultivation is required.
Addressing this request, for example, continuous cultivation of butanol-isopropyl alcohol using a microorganism separated from the soil belonging to the genus Clostridium is reported in Journal of Biochemical Engineering, 71(1), pp. 9-14, (1993). In this document, continuous cultivation is performed for 30 days. However, this microorganism is not modified by genetic recombination, and the isopropyl alcohol selection ratio thereof is as low as about 25%.
Production of isopropyl alcohol through long-term semibatch cultivation using an Escherichia coli modified to produce isopropyl alcohol is reported in J. Biosci. Bioeng., 110(6), pp. 696-701, (2010). In this document, isopropyl alcohol produced is transferred from the culture solution into the gas by gas stripping, and the isopropyl alcohol contained in the gas is recovered using water as a capturing solution. Here, an Erlenmeyer flask is used as the culture tank, and the specific surface area of the culture solution is increased by charging the culture solution in a very small amount, which is 1/10 or less of the volume of the flask; further, although the cultivation is semibatch cultivation with sequential addition, the produced isopropyl alcohol as well as the culture solution are evaporated, whereby the amount of the culture solution decreases, and a long operation time of 240 hours is possible.
J. Ind. Microbiol. Biotechnol, 33, pp. 834-844, (2006) reports ethanol continuous cultivation without aeration, in which a genetically-modified Escherichia coli for ethanol production. Here, continuous cultivation with fluid-circulation-type fixed bed is carried out in which, in addition to feeding of a sterilized culture medium to the culture tank and drawing of the culture solution from the culture tank carried out in general continuous cultivation, the solution in the culture tank is circulated.
It is known that acetic acid is produced as a by-product in aerobic cultivation using Escherichia coli in which oxygen gas or an oxygen-containing gas is supplied. An increased concentration of accumulated acetic acid causes inhibition of the growth of Escherichia coli and a decrease in the efficiency of the production of a target product. To address this issue, a DO-Stat method is well known in which the aeration or stirring rate is regulated in order to suppress high accumulation of acetic acid, and in which the concentration of dissolved oxygen in the culture tank is adjusted to several ppm in order to prevent depletion. In regard to the accumulation of acetic acid, Biotech. Bioeng., 36, pp. 750-758, (1990) reports that the concentration of acetic acid at 48 hours in the ordinary semibatch cultivation is 35 g/L, that the concentration of acetic acid at 36 hours with control by the DO-Stat method is 17 g/L, and that acetic acid is not produced in a Balanced DO-Stat method in which regulation of the concentration of glucose in the culture tank by control of the addition rate of the substrate solution in the semibatch cultivation is also carried out in addition to the regulation of the concentration of dissolved oxygen.